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Design and application of single-cell RNA sequencing to study kidney immune cells in lupus nephritis

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From Nature Reviews Nephrology

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Abstract

The immune mechanisms that cause tissue injury in lupus nephritis have been challenging to define. The advent of high-dimensional cellular analyses, such as single-cell RNA sequencing, has enabled detailed characterization of the cell populations present in small biopsy samples of kidney tissue. In parallel, the development of methods that cryopreserve kidney biopsy specimens in a manner that preserves intact, viable cells, has enabled the uniform analysis of tissue samples collected at multiple sites and across many geographic areas and demographic cohorts with high-dimensional platforms. The application of these methods to kidney biopsy samples from patients with lupus nephritis has begun to define the phenotypes of both infiltrating and resident immune cells, as well as parenchymal cells, present in nephritic kidneys. The detection of similar immune cell populations in urine suggests that it might be possible to non-invasively monitor immune activation in kidneys. Once applied to large patient cohorts, these high-dimensional studies might enable patient stratification according to patterns of immune cell activation in the kidney or identify disease features that can be used as surrogate measures of efficacy in clinical trials. Applied broadly across multiple inflammatory kidney diseases, these studies promise to enormously expand our understanding of renal inflammation in the next decade.

Key points

  • Single-cell RNA sequencing (scRNA-seq) has begun to define the phenotypes of distinct immune cell populations that accumulate in the kidney in lupus nephritis; these populations can be assessed across patients to identify molecularly distinct disease mechanisms and inform precision medicine strategies.

  • Design of cohort studies of lupus nephritis using scRNA-seq must balance the desire for optimal samples with feasibility and must consider aspects of clinical heterogeneity including disease duration, and patient gender, ethnicity and drug treatment in enrolment criteria.

  • Cryopreservation of tissue biopsies allows the accumulation of a biorepository of tissue containing intact cells, which can be analysed uniformly in batches to reduce technical variation.

  • Comparison of immune cells in the kidney with cells from urine and blood might identify cellular markers that can be measured in these biological samples reducing the need for invasive kidney biopsies.

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Fig. 1: Implementation of scRNA-seq to study lupus nephritis kidney biopsy samples.
Fig. 2: Towards a cellular classification of renal immunopathology.
Fig. 3: Insights from scRNA-seq analysis of the renal immune cell compartment in lupus nephritis.

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Acknowledgements

All authors are supported by the Accelerated Medicines Partnership (AMP) NIH UH2 AR06768. D.A.R. is supported by the Lupus Research Alliance, Burroughs Wellcome Fund Career Award for Medical Sciences and NIH NIAMS K08 AR-072791-01.

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Authors and Affiliations

  1. Division of Rheumatology, Inflammation, and Immunity, Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA, USA

    Deepak A. Rao

  2. Broad Institute of MIT and Harvard, Cambridge, MA, USA

    Arnon Arazi

  3. Rheumatology Division and Russell/Engleman Research Center, University of California San Francisco, San Francisco, CA, USA

    David Wofsy

  4. Center for Autoimmune, Musculoskeletal and Hematopoietic Diseases, The Feinstein Institute for Medical Research, Northwell Health, Manhasset, NY, USA

    Betty Diamond

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All authors researched data for the article, wrote the manuscript, made substantial contributions to discussions of the content and reviewed or edited the manuscript before submission.

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Correspondence to Betty Diamond.

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Nature Reviews Nephrology thanks T. M. Chan, V. Kyttaris and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Glossary

Unsupervised clustering

Unbiased approach used to identify sets of samples that share similar gene expression signatures.

Laser capture microscopy

A method that uses microscopy to identify and selectively collect specific regions of a tissue section for downstream analyses such as RNA expression measurement.

Computational deconvolution

In RNA sequencing analysis, an effort to quantify or identify features of individual cell populations from bulk RNA sequencing data using cell type-specific reference datasets.

Cryopreservation

The process of freezing a sample (for example, tissue or cells) for long-term storage, often using a controlled freezing method in the presence of a cryoprotectant such as dimethyl sulfoxide to preserve cell viability during freeze and thaw.

Proliferative nephritis

A form of nephritis characterized histologically by endothelial and mesangial proliferation within glomeruli.

Membranous nephritis

A form of nephritis characterized by thickening of the glomerular capillary wall with subepithelial deposits but without cellular proliferative changes.

Saturation analysis

A procedure to estimate the extent to which a given sample represents a studied population. Typically, this analysis involves down-sampling the original data, computing a statistic of interest (for example, the number of discovered clusters) and estimating its rate of change across a sequence of sample sizes.

Plasmablasts

A differentiated B cell population specialized in the secretion of antibodies; often a precursor to plasma cells.

5′ scRNA-seq

Single-cell RNA sequencing (scRNA-seq) methods that sequence RNA transcripts from the 5′ end, allowing clearer analysis of the 5′ end of RNA transcripts, as opposed to the more common strategies of RNA-seq methods that analyse transcripts from the 3′ end.

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Rao, D.A., Arazi, A., Wofsy, D. et al. Design and application of single-cell RNA sequencing to study kidney immune cells in lupus nephritis. Nat Rev Nephrol 16, 238–250 (2020). https://doi.org/10.1038/s41581-019-0232-6

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